铁皮石斛DobHLH51 基因克隆及表达特性分析

Cloning and Expression Analysis of DobHLH51 from Dendrobium officinale Kimura et Migo

bHLH转录因子(basic/helix-loop-helix)是植物响应非生物胁迫过程中重要的调控因子。为探究DobHLH51转录因子在铁皮石斛非生物胁迫响应中的表达调控机制,通过同源克隆从广东丹霞铁皮石斛叶组织中获得DobHLH51基因,并对其进行生物信息学及表达特性分析。结果显示,克隆的DobHLH51基因(Gen-Bank登录号OR234020)的CDS序列为768 bp,编码255个氨基酸,与参考序列(基因ID:LOC110107930)对比存在13个碱基差异。DobHLH51蛋白相对分子量为28.03 kDa,理论等电点为9.71,为亲水性不稳定蛋白,无信号肽和跨膜区域,含有52个磷酸化位点,具有bHLH保守结构域和ACT结构域。氨基酸序列分析表明DobHLH51与黄石斛(Dendrobium catenatum)和金钗石斛(Dendrobium nobile)的bHLH同源性最高。DobHLH51基因在广东丹霞铁皮石斛不同组织中的表达水平存在明显差异,在茎中的表达量最高。DobHLH51基因启动子序列含有多个非生物胁迫响应相关元件,且低温、干旱和ABA能够诱导DobHLH51基因显著表达。DobHLH51基因在低温和干旱胁迫下的表达模式基本一致,但其在ABA处理下的表达模式不同,推测该基因可能通过非依赖于ABA的信号转导途径参与低温和干旱胁迫响应过程。本研究结果可为后续进一步探究DobHLH51基因功能及其在非生物胁迫响应中的作用机制提供理论依据。

Basic/helix-loop-helix(bHLH)transcription factor is a key regulatory factor in plant abiotic stress responses.In order to explore the regulatory mechanism of DobHLH51 from Dendrobium officinale in re-sponse to abiotic stress,the DobHLH51 gene was cloned from leaves using homologous cloning method,and its bioinformatics and expression characteristics were analyzed.The results showed that the full-length coding se-quence(CDS)of DobHLH51 was 768 bp and encoded 255 amino acids,which had 13 bases differences com-pared with the reference sequence(Gene ID:LOC110107930).The theoretical molecular weight and isoelec-tric point of DobHLH51 protein were 28.03 kDa and 9.71,respectively.It was a hydrophilic and unstable pro-tein with no signal peptide or transmembrane region,which contained 52 phosphorylation sites and possessed a bHLH conserved domain and an ACT domain.Results of amino acid sequence analysis showed that DobHLH51 had the highest homology with other bHLH proteins in D.catenatum and D.nobile.The expressions of DobHLH51 showed obvious differences among tissues of D.officinale from Danxia of Guangdong,which were the highest in stems.The DobHLH51 promoter region contained several cis-acting elements in response to abi-otic stresses,and its expression could be significantly induced by cold,drought stress and ABA treatment.The expression pattern of DobHLH51 was basically consistent under cold and drought stresses,which was different when treated with ABA,indicating that DobHLH51 might participate in cold and drought stress responses through a transduction pathway independent of ABA.This study could provide theoretical bases for further re-search on the function of DobHLH51 and its role in abiotic stress response.